ESEEM Reveals Bound Substrate Histidine in the ABC Transporter HisQMP 2

Nikolay Isaev, Johanna Heuveling, Nikita Ivanisenko, Erwin Schneider, Heinz Jürgen Steinhoff

Research output: Contribution to journalArticlepeer-review

Abstract

Localization of substrates in membrane proteins is an important but challenging task. In this paper, we show that deuterium electron spin echo envelope modulation spectroscopy ( 2 H ESEEM) combined with site-directed spin labeling is a powerful tool to localize the substrate, histidine-d5, in the ABC transporter HisQMP 2 . Based on a homology model and spin label rotamer analyses, we calculated 2 H ESEEM spectra for eight possible labeling positions close to the putative substrate-binding site. Experimental 2 H ESEEM spectra were determined with spin labels bound either at position 169 of HisM, for which a detectable 2 H ESEEM signal was calculated, or with a spin label bound at position 54 of HisQ as a negative control. The agreement between the calculated and experimental ESEEM spectra provides strong evidence for the histidine located in a binding site primarily liganded by residues of HisM as proposed by the homology model.

Original languageEnglish
Pages (from-to)883-893
Number of pages11
JournalApplied Magnetic Resonance
Volume50
Issue number7
DOIs
Publication statusPublished - 1 Jul 2019

Keywords

  • ELECTRON-SPIN-ECHO
  • ENVELOPE MODULATION
  • WATER CONCENTRATION
  • EPR SPECTROSCOPY
  • MEMBRANE
  • BINDING
  • RADICALS
  • INVESTIGATE
  • DIVERSITY
  • PROTEINS

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